1. Field of the Invention.
The present invention relates to a flow cytometry apparatus, and more particularly, concerns a flow cytometry apparatus for determining one or more characteristics of cells or the like which includes improved optics for the light beam adjustment.
2. Description of the Prior Art.
Flow analysis of particles, such as cells, has been employed in the determination of different characteristics of those particles. Flow cytometry apparatuses have long been utilized for this purpose. In the broadest sense, a flow cytometry apparatus as used and meant herein is an instrument which analyzes cells or particles as they serially flow, substantially one at a time, through a sensing region. Cell volume, size, shape and identification are parameters which are typically determined in a flow cytometry apparatus, particularly as such parameters are related to a source of light directed at the cells when flowing through the sensing region. Light scattered by the flowing cells may be detected at variety of angles with respect to the axis of the incident light beam. Scattered light has served as a function of cell shape, index of refraction, opacity, roughness and the like. Fluorescence emitted by labeled cells which have been excited as a result of passing through the excitation energy of the incident light beam is detectable for identification of specifically labeled cells. Not only is cell analysis performed on the flow cytometry apparatuses, but sorting of cells may also be achieved. In U.S. Pat. No. 3,826,364, a flow cytometry apparatus is disclosed which physically separates particles such as functionally different cell types. In this patented cell sorter, a laser provides illumination which is focused on the stream of particles by a suitable lens or lens system so that there is highly localized scatter from particles therein. In addition, a high intensity source of illumination is directed onto the stream of particles for the excitation of fluorescent particles in the stream. Certain particles in the stream may be selectively charged and then separated by deflecting them into designated receptacles.
When utilizing lasers or other light sources for illumination in flow cytometry apparatuses, obtaining optimum fluorescent pulse height resolution involves a balance between illumination uniformity, which determines the uniformity of fluorescence with particle position, and laser beam intensity, which determines the available fluorescence photon flux. One such technique of improving fluorescence sensitivity in the flow cytometry apparatus is disclosed in U.S. Pat. No. 4,498,766. In the aforementioned patent, fluorescence sensitivity is improved by positioning the focusing lens at an angle relative to the light beam so that the light beam focal waist becomes elongated, and elliptical in nature. The elliptical focal spot allows the light energy from the laser to be focused into a focal spot so that the energy distribution in the direction of particle travel is optimized thereby affecting fluorescence sensitivity. Other light beam focusing devices are described in U.S. Pat. Nos. 4,293,221; 4,243,318 and 3,606,547.
Even though the focused laser or other light beam provides the improvement in fluorescence sensitivity and optimization of the focal spot intensity, optimal performance is minimized if the focused light beam is not properly adjusted on the stream of flowing cells Many flow cytometry apparatuses include one or more devices for adjusting the positioning of the focused light beam on the flowing cells. Conventional positioning methods employ expensive differential micrometers to position the light source itself, or might include optical elements such as mirrors or prisms. Since the biological cells or particles under analysis are typically a few microns in size, the precision of the light beam adjustment is also in the micron range, thus requiring high resolution mechanical displacement devices Simple and inexpensive techniques of adjusting or positioning a focused light beam on the flowing cells are still being sought. The present invention is directed to the discovery of a straightforward mechanism to obtain fine position adjustment of the focused light beam on the cells.